Hepatitis C virus (HCV) infects approximately 1% of the world's population and causes serious health problems. Over 75% of acutely infected individuals eventually progress to a chronic carrier state that can result in cirrhosis, liver failure, and hepatocellular carcinoma. A very small fraction of chronically infected patients clear HCV naturally and resolve chronic hepatitis. See Alter et al. (1992) N. Engl. J. Med. 327:1899-1905; Resnick and Koff. (1993) Arch. Intem. Med. 153:1672-1677; Seeff (1995) Gastrointest. Dis. 6:20-27; Tong et al. (1995) N. Engl. J. Med. 332:1463-1466. Immunization against E2 glycoproteins of some flaviviruses (see e.g., Konishi et al., (1992) Virology 188: 714-720), including HCV (Ishii et al., (1998) Hepatology 28: 1117-1120), may protect against infection. However, attempts to express recombinant HCV E1 and E2 glyocoproteins have been frustrated by the fact that these proteins are not secreted from the host cell but are retained within the endoplasmic reticulum (Dubuisson et al. (1994) J. Virology 68: 6147-6160).
One approach to making vaccines for HCV and other viruses which has been attempted is to prepare chimeric antigens consisting of fusions of hepatitis B virus surface antigen (HBsAg) with a heterologous antigen, for example a portion of an HCV protein. See, e.g., Inchauspe et al. (1998) Dev. Biol. Stand. 92: 162-168; Nakano et al. (1997) J. Virol. (1997) 71: 7101-7109; and Inchauspe et al. (1997) Vaccine 15: 853-856. The use of HBsAg is attractive for the production of immunogenic compositions such as vaccines because HBsAg is highly immunogenic and is secreted from cultured cells in the form of virus-like particles (U.S. Pat. No. 5,098,704). Attempts to introduce small portions of viral proteins into HBsAg have succeeded in the production of virus-like particles (see e.g., Delpeyroux et al. (1990) J. Virology 64: 6090-6100, who inserted an 11 amino acid segment of polio virus capsid protein into HBsAg). However, in one study only two out of six fusion proteins containing HBsAg combined with different hydrophillic domains of HCV E2 were secreted into the culture medium as virus-like particles (Lee et al. (1996) J. Med. Virol. 50: 145-151), possibly because the E2 inserts were too large or hydrophilic. The insertion site of heterologous epitopes into HBsAg may be an important factor. A study which inserted an epitope of HBV nucleocapsid (HBcAg) at various positions into HBsAg found that insertion into an internal site in HBsAg resulted in a chimeric protein that was immunogenic for HBcAg, while insertion at the C-terminus was weakly immunogenic (Schodel et al. (1992) J. Virology 66: 106-114). Insertion at the N-terminus prevented surface access of the HBcAg epitope in the resultant particles and was non-immunogenic (Id.). Apparently, the molecular context in which an epitope is presented is important in determining immunogenicity, probably because of subtle alterations of protein secondary and tertiary structure. This principle was further illustrated by Eckhart et al. ((1996) J. Gen. Virol. 77: 2001-2008), who introduced a conserved, six amino acid epitope of HIV-1 gp41 protein into influenza hemagglutinin and obtained neutralizing antibodies, but could not generate neutralizing antibodies when the same epitope was inserted into HBsAg. Smaller isolated epitopes are more likely to be sensitive to such effects than larger portions of an immunogenic protein.
Currently there is no method available for expressing entire E1 or E2 glycoproteins of HCV in virus-like particles for use in immunization. Available methods limit chimeric proteins based on HBsAg to the insertion of only small isolated domains of E2, which may or may not have a native immunogenic structure. Thus, there remains a need in the art for methods and materials that can be used to express HCV antigens in an immunogenic form in virus-like particles.